New Advances in Dye Analyses: In Situ Gel-Supported Liquid Extraction from Paint Layers and Textiles for SERS and HPLC-MS/MS Identification.

To date, it is still not possible to obtain exhaustive information about organic materials in cultural heritage without sampling. Nonetheless, when studying unique objects with invaluable artistic or historical significance, preserving their integrity is a priority. In particular, organic dye identification is of significant interest for history and conservation research, but it is still hindered by analytes' low concentration and poor fastness. In this work, a minimally invasive approach for dye identification is presented. The procedure is designed to accompany noninvasive analyses of inorganic substances for comprehensive studies of complex cultural heritage matrices, in compliance with their soundness. Liquid extraction of madder, turmeric, and indigo dyes was performed directly from paint layers and textiles. The extraction was supported by hydrogels, which themselves can undergo multitechnique analyses in the place of samples. After extraction, Ag colloid pastes were applied on the gels for SERS analyses, allowing for the identification of the three dyes. For the HPLC-MS/MS analyses, re-extraction of the dyes was followed by a clean-up step that was successfully applied on madder and turmeric. The colour change perceptivity after extraction was measured with colorimetry. The results showed ΔE values mostly below the upper limit of rigorous colour change, confirming the gentleness of the procedure.

Inside the History of Italian Coloring Industries: An Investigation of ACNA Dyes through a Novel Analytical Protocol for Synthetic Dye Extraction and Characterization.

The introduction of synthetic dyes completely changed the industrial production and use of colorants for art materials. From the synthesis of the first synthetic dye, mauveine, in 1856 until today, artists have enjoyed a wider range of colors and selection of chemical properties than was ever available before. However, the introduction of synthetic dyes introduced a wider variety and increased the complexity of the chemical structures of marketed dyes. This work looks towards the analysis of synthetically dyed objects in heritage collections, applying an extraction protocol based on the use of ammonia, which is considered favorable for natural anthraquinone dyes but has never before been applied to acid synthetic dyes. This work also presents an innovative cleanup step based on the use of an ion pair dispersive liquid-liquid microextraction for the purification and preconcentration of historical synthetic dyes before analysis. This approach was adapted from food science analysis and is applied to synthetic dyes in heritage science for the first time in this paper. The results showed adequate recovery of analytes and allowed for the ammonia-based extraction method to be applied successfully to 15 samples of suspected azo dyes from the Azienda Coloranti Nazionali e Affini (ACNA) synthetic dye collection, identified through untargeted HPLC-HRMS analyses.

Smartphone-Based Electrochemical Biosensor for On-Site Nutritional Quality Assessment of Coffee Blends.

This work aimed to develop an easy-to-use smartphone-based electrochemical biosensor to quickly assess a coffee blend's total polyphenols (Phs) content at the industrial and individual levels. The device is based on a commercial carbon-based screen-printed electrode (SPE) modified with multi-walled carbon nanotubes (CNTs) and gold nanoparticles (GNPs). At the same time, the biological recognition element, Laccase from Trametes versicolor, TvLac, was immobilized on the sensor surface by using glutaraldehyde (GA) as a cross-linking agent. The platform was electrochemically characterized to ascertain the influence of the SPE surface modification on its performance. The working electrode (WE) surface morphology characterization was obtained by scanning electron microscopy (SEM) and Fourier-transform infrared (FT-IR) imaging. All the measurements were carried out with a micro-potentiostat, the Sensit Smart by PalmSens, connected to a smartphone. The developed biosensor provided a sensitivity of 0.12 µA/µM, a linear response ranging from 5 to 70 µM, and a lower detection limit (LOD) of 2.99 µM. Afterward, the biosensor was tested for quantifying the total Phs content in coffee blends, evaluating the influence of both the variety and the roasting degree. The smartphone-based electrochemical biosensor's performance was validated through the Folin-Ciocâlteu standard method.

A review on inorganic gaseous pollutants in conservation spaces: monitoring instrumentation and indoor concentrations.

This contribution presents the results of a review of scientific literature on gaseous inorganic pollutants monitored in confined indoor spaces housing cultural heritage. A survey on standards suggesting concentration thresholds together with European projects on the topic was provided. Sixty-six scientific articles were systematically selected based on the PRISMA flow diagram over the period 1984-2021 for a total number of 80 case studies mainly located in Europe (64%). Monitoring was mainly performed in museums and galleries (61%), specifically in exhibition rooms (79%). Active devices were rarely employed, whereas passive samplers, exposed in situ and then laboratory-analysed, were mostly used for nitrogen dioxide and sulphur dioxide monitoring. Direct-reading continuous devices were widely used for ozone monitoring. It was found that average concentrations of ozone were below 5 ppb in only 50% of cases, nitrogen dioxide below 10 ppb in more than 60% of cases, nitric oxide below 5 ppb in 30% of cases, nitric and nitrous acid below 1 ppb in less than 50% of cases, sulphur dioxide below 2 ppb in more than 60% of cases, and hydrogen sulphide below 0.1 ppb in only 25% of cases. Comparisons were performed following the thresholds suggested in the literature. The lowest concentration values were usually associated to the use of mechanical ventilation systems equipped with air filters and to non-urban case studies. The low number of case studies can be due to the difficulties to perform monitoring in conservation spaces with current instruments. Further research should be conducted to uniform standards that suggest instruments' requirements and pollutant thresholds to limit degradation on cultural materials.

Label-free magnetic nanoparticles-based electrochemical immunosensor for atrazine detection.

This work presents the realization of a label-free electrochemical immunosensor for the quick, cheap, and straightforward determination of atrazine. This biodevice is based on developing a technological platform where a gold screen printed electrode (Au-SPE) surface was modified by the electrodeposition of a highly porous gold layer. As an internal probe redox, a Prussian Blue thin layer (PB) was then electrosynthetized onto the modified Au-SPE. Atrazine antibody (Ab-ATZ) was immobilized using G protein-functionalized magnetic nanoparticles (MNPs@protG) to ensure the correct orientation of the antibody to enhance the immunoaffinity. Under optimum experimental conditions, the electrochemical characterization of the developed immunosensor displays a linearity range towards atrazine within 0.05-1.5 ng/mL, a LOD of 0.011 ng/mL good reproducibility and stability. The immunosensor was tested in the analysis of spiked drinking water samples with a mean recovery ranging from 95.7 to 108.4%. The overall good analytical performances of this immunodevice suggest its application for the screening and monitoring of atrazine in real matrices.

Phthalate esters (PAEs) concentration pattern reflects dietary habitats (δ13C) in blood of Mediterranean loggerhead turtles (Caretta caretta).

Phthalic acid esters (PAEs) are classified as endocrine disruptors, but it remains unclear if they can enter the marine food-web and result in severe health effects for organisms. Loggerhead turtles (Caretta caretta) can be chronically exposed to PAEs by ingesting plastic debris, but no information is available about PAEs levels in blood, and how these concentrations are related to diet during different life stages. This paper investigated, for the first time, six PAEs in blood of 18 wild-caught Mediterranean loggerhead turtles throughout solid-phase extraction coupled with gas chromatography-ion trap/mass spectrometry. Stable isotope analyses of carbon and nitrogen were also performed to assess the resource use pattern of loggerhead turtles. DEHP (12-63 ng mL-1) and DBP (6-57 ng mL-1) were the most frequently represented PAEs, followed by DiBP, DMP, DEP and DOP. The total PAEs concentration was highest in three turtles (124-260 ng mL-1) whereas three other turtles had concentrations below the detection limit. PAEs were clustered in three groups according to concentration in all samples: DEHP in the first group, DBP, DEP, and DiBP in the second group, and DOP and DMP in the third group. The total phthalates concentration did not differ between large-sized (96.3 ± 86.0 ng mL-1) and small-sized (67.1 ± 34.2 ng mL-1) turtles (p < 0.001). However, DMP and DEP were found only in large-sized turtles and DiBP and DBP had higher concentrations in large-sized turtles. On the other hand, DEHP and DOP were found in both small- and large-sized turtles with similar concentrations, i.e. ~ 21.0/32.0 ng mL-1 and ~ 7.1/9.9 ng mL-1, respectively. Winsored robust models indicated that δ13C is a good predictor for DBP and DiBP concentrations (significant Akaike Information criterion weight, AICwt). Our results indicate that blood is a good matrix to evaluate acute exposure to PAEs in marine turtles. Moreover, this approach is here suggested as a useful tool to explain the internal dose of PAEs in term of dietary habits (δ13C), suggesting that all marine species at high trophic levels may be particularly exposed to PAEs, despite their different dietary habitats and levels of exposure.

Photons for Photography: A First Diagnostic Approach to Polaroid Emulsion Transfer on Paper in Paolo Gioli's Artworks.

The aim of this research is to study and diagnose for the first time the Polaroid emulsion transfer in the contemporary artist Paolo Gioli's artworks to provide preliminary knowledge about the materials of his artworks and the appropriate protocols which can be applied for future studies. The spectral analysis performed followed a multi-technical approach first on the mock-up samples created following Gioli's technique and on one original artwork of Gioli, composed by: FORS (Fiber Optics Reflectance), Raman, and FTIR (Fourier-Transform InfraRed) spectroscopies. These techniques were chosen according to their completely non-invasiveness and no requirement for sample collection. The obtained spectra from FTIR were not sufficient to assign the dyes found in the transferred Polaroid emulsion. However, they provided significant information about the cellulose-based materials. The most diagnostic results were obtained from FORS for the determination of the dye developers present in the mock-up sample which was obtained from Polacolor Type 88 and from Paolo Gioli's original artwork created with Polacolor type 89.

Single-Sided Portable NMR Investigation to Assess and Monitor Cleaning Action of PVA-Borax Hydrogel in Travertine and Lecce Stone.

In this work, we investigated the potential of PVA-borax hydrogel for cleaning limestones and the dependence of the cleaning on the porosity of the rock and on the action time of the hydrogel treatment. Towards this goal, we used a nuclear magnetic resonance (NMR) spectrometer, developed for non-invasive and non-destructive applications on cultural heritage. T2-NMR parameters were quantified on different samples of Lecce stone and Travertine cut perpendicular (Pe) and parallel (Pa) to the bedding planes under different experimental conditions: untreated samples, treated with Paraloid B72 and cleaned with PVA-PEO-borax hydrogel applied for 4 min and 2 h. The T2 results suggest that the effectiveness of the cleaning strongly depended on the porosity of the stones. In Lecce stone, the hydrogel seemed to eliminate both the paramagnetic impurities (in equal measure with 4 min and 2 h treatment) and Paraloid B72. In Travertine Pe, characterized by a smaller pore size compared to Lecce stone, no significant effects were found regarding both the cleaning and the treatment with Paraloid B72. In Travertine Pa, characterized by a larger pore size than the other two samples, the hydrogel seemed to clean the paramagnetic agents (it worked better if applied for a longer time) but it did not appear to have any effect on Paraloid B72 removal.

Site-Directed Antibody Immobilization by Resorc[4]arene-Based Immunosensors.

One of the main problems in the development of immunosensors is to overcome the complexity of binding antibodies to the sensor surface. Most immobilizing methods lead to a random orientation of antibodies with a lower binding site density and immunoaffinity. In order to control the orientation of antibody immobilization, several resorc[4]arene derivatives were designed and synthesized. After the spectroscopic characterization of resorc[4]arene self-assembled monolayers (SAMs) onto gold films, the surface coverage and the orientation of insulin antibody (Ab-Ins) were assessed by a surface plasmon resonance (SPR) technique and compared with a random immobilization method. Experimental results combined with theoretical studies confirmed the dipole-dipole interaction as an important factor in antibody orientation and demonstrated the importance of the upper rim functionalization of resorcarenes. Accordingly, resorcarene 5 showed a major binding force towards Ab-Ins thanks to the H-bond interactions with the amine protein groups. Based on these findings, the resorcarene-based immunosensor is a powerful system with improved sensitivity providing new insight into sensor development.

A bimetallic nanocoral Au decorated with Pt nanoflowers (bio)sensor for H2O2 detection at low potential.

In this work, we have developed for the first time a method to make novel gold and platinum hybrid bimetallic nanostructures differing in shape and size. Au-Pt nanostructures were prepared by electrodeposition in two simple steps. The first step consists of the electrodeposition of nanocoral Au onto a gold substrate using hydrogen as a dynamic template in an ammonium chloride solution. After that, the Pt nanostructures were deposited onto the nanocoral Au organized in pores. Using Pt (II) and Pt (IV), we realized nanocoral Au decorated with Pt nanospheres and nanocoral Au decorated with Pt nanoflowers, respectively. The bimetallic nanostructures showed better capability to electrochemically oxidize hydrogen peroxide compared with nanocoral Au. Moreover, Au-Pt nanostructures were able to lower the potential of detection and a higher performance was obtained at a low applied potential. Then, glucose oxidase was immobilized onto the bimetallic Au-Pt nanostructure using cross-linking with glutaraldehyde. The biosensor was characterized by chronoamperometry at +0.15V vs. Ag pseudo-reference electrode (PRE) and showed good analytical performances with a linear range from 0.01 to 2.00mM and a sensitivity of 33.66µA/mMcm2. The good value of Kmapp (2.28mM) demonstrates that the hybrid nanostructure is a favorable environment for the enzyme. Moreover, the low working potential can minimize the interference from ascorbic acid and uric acid as well as reducing power consumption to effect sensing. The simple procedure to realize this nanostructure and to immobilize enzymes, as well as the analytical performances of the resulting devices, encourage the use of this technology for the development of biosensors for clinical analysis.

Comparison between a Direct-Flow SPR Immunosensor for Ampicillin and a Competitive Conventional Amperometric Device: Analytical Features and Possible Applications to Real Samples.

In this research, we developed a direct-flow surface plasmon resonance (SPR) immunosensor for ampicillin to perform direct, simple, and fast measurements of this important antibiotic. In order to better evaluate the performance, it was compared with a conventional amperometric immunosensor, working with a competitive format with the aim of finding out experimental real advantages and disadvantages of two respective methods. Results showed that certain analytical features of the new SPR immunodevice, such as the lower limit of detection (LOD) value and the width of the linear range, are poorer than those of a conventional amperometric immunosensor, which adversely affects the application to samples such as natural waters. On the other hand, the SPR immunosensor was more selective to ampicillin, and measurements were more easily and quickly attained compared to those performed with the conventional competitive immunosensor.

Inhibition-based first-generation electrochemical biosensors: theoretical aspects and application to 2,4-dichlorophenoxy acetic acid detection.

In this work, several theoretical aspects involved in the first-generation inhibition-based electrochemical biosensor measurements have been discussed. In particular, we have developed a theoretical-methodological approach for the characterization of the kinetic interaction between alkaline phosphatase (AlP) and 2,4-dichlorophenoxy acetic acid (2,4-D) as representative inhibitor studied by means of cyclic voltammetry and amperometry. Based on these findings, a biosensor for the fast, simple, and inexpensive determination of 2,4-D has been developed. The enzyme has been immobilized on screen-printed electrodes (SPEs). To optimize the biosensor performances, several carbon-based SPEs, namely graphite (G), graphene (GP), and multiwalled carbon nanotubes (MWCNTs), have been evaluated. AlP was immobilized on the electrode surface by means of polyvinyl alcohol with styryl-pyridinium groups (PVA-SbQ) as cross-linking agent. In the presence of ascorbate 2-phosphate (A2P) as substrate, the herbicide has been determined, thanks to its inhibition activity towards the enzyme catalyzing the oxidation of A2P to ascorbic acid (AA). Under optimum experimental conditions, the best performance in terms of catalytic efficiency has been demonstrated by MWCNTs SPE-based biosensor. The inhibition biosensor shows a linearity range towards 2,4-D within 2.1-110 ppb, a LOD of 1 ppb, and acceptable repeatability and stability. This analysis method was applied to fortified lake water samples with recoveries above 90%. The low cost of this device and its good analytical performances suggest its application for the screening and monitoring of 2,4-D in real matrices.

Amine oxidase-based biosensors for spermine and spermidine determination.

The present work describes the development and optimization of electrochemical biosensors for specific determination of the biogenic polyamine spermine (Spm) and spermidine (Spmd) whose assessment represents a novel important analytical tool in food analysis and human diagnostics. These biosensors have been prepared using novel engineered enzymes: polyamine oxidase (PAO) endowed with selectivity towards Spm and Spmd and spermine oxidase (SMO) characterized by strict specificity towards Spm. The current design entails biosensors in which the enzymes were entrapped in poly(vinyl alcohol) bearing styrylpyridinium groups (PVA-SbQ), a photocrosslinkable gel, onto an electrode surface. Screen-printed electrodes (SPEs) were used as electrochemical transducers for enzymatically produced hydrogen peroxide, operating at different potential vs Ag/AgCl according to the material of the working electrode (WE): +700 mV for graphite (GP) or -100 mV for Prussian blue (PB)-modified SPE, respectively. Biosensor performances were evaluated by means of flow injection amperometric (FIA) measurements. The modified electrodes showed good sensitivity, long-term stability and reproducibility. Under optimal conditions, the PAO biosensor showed a linear range 0.003-0.3 mM for Spm and 0.01-0.4 mM for Spmd, while with the SMO biosensor, a linear range of 0.004-0.5 mM for Spm has been obtained. The main kinetic parameters apparent Michaelis constant (K M), turnover number (K cat) and steady-state current (I max) were determined. The proposed device was then applied to the determination of biogenic amines in blood samples. The results obtained were in good agreement with those obtained with the GC-MS reference method.

Development of Carbon-Based Nano-Composite Materials for Direct Electron Transfer Based Biosensors.

Nafion, an ion exchange polymer that is very resistant to chemical attack, even by strong oxidant at high temperatures, has found great increasing use as a film material; however, its use as immobilizing agent in third-generation biosensors is hindered due to the low rate of charge transfer in the pure Nafion film. In this work we showed that the use of functionalized multi-walled carbon nanotubes Nafion/MWCNTs composite film for modification of the carbon-based electrode surfaces would increase the charge transfer rate greatly; the composite has proven to efficiently immobilize two different heme proteins (catalase and cytochrome c) and to enhance the electrochemical performances of several carbon electrode materials (glassy carbon, mesoporous graphite, graphite and graphene) either used as classical electrodes or screen printed ones. The electrochemical signal of both redox proteins becomes more reversible and the electron transfer kinetic constant increases. At the same time the biological activity is maintained indicating that the immobilization procedure allows the proteins to retain a native-like structure.

A new surface plasmon resonance immunosensor for triazine pesticide determination in bovine milk: a comparison with conventional amperometric and screen-printed immunodevices.

A detailed comparison was made of the analytical features of a new Surface Plasmon Resonance (SPR) immunodevice for triazine pesticide determination with those of two other amperometric (conventional and screen-printed) immunosensors and the advantages and disadvantages of the SPR method were thoroughly investigated. For conventional amperometric and screen-printed devices, "competitive" assays were used; conversely, the SPR transduction technique allowed a "direct" measurement format to be used. As far as the main analytical data are concerned, the SPR method does not seem to offer substantial advantages. Nevertheless the measurement time is much shorter and the measurement itself much easier to perform. Lastly several applications and recovery tests were carried out on bovine milk samples, before and after spiking, to check for triazine pesticides in the samples, obtaining satisfactory results.

Multi-Technique Assessment of Chelators-Loaded PVA-Borax Gel-like Systems Performance in Cleaning of Stone Contaminated with Copper Corrosion Products.

Currently, one of the most important challenges for the conservation of stone artworks is the removal of metal corrosion products on their surfaces. Traditional cleaning methods, which typically involve the application of aqueous solutions containing chelating agents capable of complexing these metal ions, have shown some weaknesses. These weaknesses become apparent when such methods are applied to statues and other vertical surfaces or when aiming to limit the cleaning process to a specific area with controlled application times. Furthermore, the porosity of the stone surface plays a role concerning the cleaning efficiency. To address these issues, chelating agents can be incorporated into gel-like materials. This study is a proof of concept to evaluate the cleaning efficacy of various gel formulations composed of polyvinyl alcohol (PVA), borax (B), and agarose (AG), loaded with two chelators: ethylenediaminetetraacetic acid (EDTA) and potassium sodium tartrate (PST or Rochelle salt). Three types of carbonate stones (travertine, Lecce stone, and Carrara marble) characterized by different porosities were artificially stained with copper sulphates and treated with the different PVA-B-AG formulations. The effectiveness of the treatment was directly monitored on the stones using a multi-technique approach that included scanning electron microscopy with energy dispersive spectroscopy (SEM-EDS) and non-invasive portable nuclear magnetic resonance (NMR). Additionally, the rheological properties of the gels were investigated, and the Fourier transform infrared attenuated total reflection spectroscopy (FTIR ATR) was used to analyse the chemical structure of the gel before and after treatment, aiming to understand the changes induced by the cleaning process.

Early Stages of Metal Corrosion in Coastal Archaeological Sites: Effects of Chemical Composition in Silver and Copper Alloys.

In this study, metal disks with different chemical composition (two Ag-based alloys and three Cu-based alloys) were buried in the soil of coastal archaeological sites for a period of 15 years. The aim was to naturally induce the growth of corrosion patinas to obtain a deeper insight into the role of alloying elements in the formation of the patinas and into the degradation mechanisms occurring in the very early stages of burial. To reach the aim, the morphological, compositional and structural features of the patinas grown over 15 years were extensively characterized by optical microscopy, field emission scanning electron microscopy coupled with energy dispersive spectrometry, X-ray diffraction and micro-Raman spectroscopy. Results showed that the Cu amount in Ag-based alloys strongly affected the final appearance, as well as the composition and structure of the patinas. Corrosion mechanisms typical of archaeological finds, such as the selective dissolution of Cu, Pb and Zn and internal oxidation of Sn, occurred in the Cu-based alloys, even if areas enriched in Zn and Pb compounds were also detected and attributed to an early stage of degradation. In addition, some unusual and rare compounds were detected in the patinas developed on the Cu-based disks.

Natural based products for cleaning copper and copper alloys artefacts.

Copper alloys objects can deteriorate their conservation state through irreversible corrosion. Since in the cultural heritage field every artefact is unique and any loss irreplaceable, solutions for conservation are needed. Hence, there is the necessity to stop the corrosion process with a suitable cleaning and conservation process to avoid further degradation processes without changing its morphological aspect. Chelating solutions are commonly used in chemical cleaning, mainly sodium salts of ethylenediaminetetraacetic acid (EDTA). However, it is resistant to water purification procedures and is not biodegradable. The goal of this study was to see if applying an ecologically friendly chelating agent as an alternative to EDTA cleaning procedures for cultural heritage was suitable. In this study were chosen six natural-based chelators that could be a new green non-toxic alternative to EDTA in corrosion-inhibiting properties. They were tested for cleaning copper artefacts exposed to atmospheric environment in polluted areas. The study considered four amino acids, a glucoheptonate (CSA) and an industrial green chelator (GLDA). The effectiveness was tested on corrosion copper compounds and on laboratory corroded copper sheets. Finally, the cleaning efficacy was tested on four Roman coins and a modern copper painting. To define the cleaning efficacy, surface analytical investigations have been carried out by means ICP-OES, UV-VIS, µ-Raman, spectro-colorimetry, XRD and FTIR. Among the amino acids, alanine was the most effective, showing an unaltered noble patina and a good effective copper recovery from corrosion patinas.

ASu@MNPs-based electrochemical immunosensor for vitamin D3 serum samples analysis.

We report a new sensitive label-free electrochemical immunosensor to detect Vitamin D3 (25-OHD3) in untreated serum samples. To this aim, a graphite screen printed electrode (SPE) was modified using cysteamine (CYM) functionalized core-shell magnetic nanoparticles (Au@MNPs) then, the 25-OHD3 antibody (AbD) was immobilized via glutaraldehyde crosslinking. The several steps involved in the immunosensor development and 25-OHD3 analysis were monitored by using differential pulse voltammetry (DPV). The developed immunosensor showed a LOD of 2.4 ng mL-1 and a linear range between 7.4 and 70 ng mL-1. The effectiveness of the immunosensor in human serum analysis was assessed by comparing the results obtained with the chemiluminescence-immunoassay (CLIA) reference method. The high sensitivity and excellent agreement with the reference method suggest its potential use as a POCT to monitor hypovitaminosis 25-OHD levels.

Applying Gel-Supported Liquid Extraction to Tutankhamun's Textiles for the Identification of Ancient Colorants: A Case Study.

The identification of the dyes present on a linen fragment from the tomb of Pharaoh Tutankhamun is the objective of the present study. Fiber optic reflectance spectroscopy (FORS) was applied to the archaeological sample for preliminary identification of the dyes and to better choose the extraction methodology for different areas of the sample. The innovative gel-supported micro-extraction with agar gel and the Nanorestore Gel® High Water Retention (HWR) gel were applied to the archaeological sample after testing of the best concentration for the extraction of the agar gels substrates, performed on laboratory mock-ups by means of UV-Vis transmittance spectroscopy. Immediately after extraction, Ag colloidal pastes were applied on the gel surface and Surface Enhanced Raman Scattering (SERS) analysis was performed directly on them. The combination of information deriving from FORS and SERS spectra resulted in the successful identification of both indigo and madder and, in hypothesis, of their degradation products.